Summary: To determine whether IgG subclasses differ in their ability to neutralize HIV-1, polyclonal HIVIG was passed over protein A columns and the appropriate fractions collected and shown to be > 89% pure IgG1, IgG2 and IgG3 (IgG4 was not present in sufficient quantities for study). All three subclasses showed binding to all major HIV-1 proteins (IgG1 >IgG2 > IgG3). In contrast, IgG3 was more active than other subclasses in its ability to neutralize HIV-1 as assayed by syncytia inhibition and cell-free virus neutralization (IgG3 > IgG1 > IgG2). The ability of HIVIG and the subclasses to neutralize viruses also depended on the virus strain, so that T cell tropic (R4) were more susceptible than Macrophage tropic (R5) viruses. IgG3 differs from IgG1 and IgG2 by virtue of a longer and more flexible hinge region. To test whether this is important in neutralizing virus, the IgG fractions were digested by papain into Fc and Fab fragments and the Fab fragments tested for activity. The hinge region is present in the F(ab)2, but not in the Fab fragments. F(ab)2, but not Fab, fragments from IgG3, retained their superior ability to neutralize HIV-1 when compared to F(ab)2 fragments from other IgG subclasses, indicating that the longer hinge region of IgG3 enables IgG3 to neutralize HIV-1 more efficiently. These findings have implications for manufacture and clinical use of HIVIG, particularly since IgG3 is more vulnerable to proteolytic enzymes than other isotypes. Moreover they suggest that HIV-1 vaccines should induce high titer IgG3 responses.